目的 探究芒柄花素磺酸鈉（Sul-F）調(diào)節(jié)中性粒細(xì)胞胞外誘捕網(wǎng)（NETs）減輕腦缺血再灌注（I/R）損傷的作用機制。方法 SD大鼠隨機分為5組：對照組、模型組、精氨酸阿司匹林組（AAFI，200 mg·kg^-1）和Sul-F低、高劑量組（20、40 mg·kg^-1）。除對照組外，其余各組均采用大腦中動脈栓塞（MCAO）法造模，缺血2 h、再灌注24 h后處死。于再灌注0、12 h后ip給藥，對照組和模型組給予等體積0.9%氯化鈉溶液。Bederson評分法評估神經(jīng)功能；TTC染色法測定腦梗死率；Western blotting檢測腦勻漿磷脂酶C-β3（PLC β3）和磷酸化PLC β3（p-PLC β3）的表達；酶聯(lián)免疫法檢測腦勻漿血栓素B2（TXB2）及炎癥因子白細(xì)胞介素-1β（IL-1β）、白細(xì)胞介素-6（IL-6）含量；免疫熒光檢測血小板活化相關(guān)分子P選擇素（Pselectin）、中性粒細(xì)胞活化相關(guān)分子中性粒細(xì)胞表面黏附分子（CD11b）的表達；三色免疫熒光聯(lián)合檢測組蛋白3B（H3B）、髓過氧化物酶（MPO）和脫氧核糖核酸（DNA），以標(biāo)識NETs。結(jié)果 與對照組相比，模型組大鼠神經(jīng)功能評分、腦指數(shù)、腦梗死率以及腦勻漿IL-1β、IL-6、TXB2蛋白表達顯著升高（P＜0.01），p-PLC β3蛋白表達顯著降低（P＜0.01），大量NETs形成，P-selectin和CD11b表達均升高（P＜0.01）。與模型組比較，Sul-F顯著降低神經(jīng)功能評分、腦指數(shù)、腦梗死率以及腦勻漿IL-1β、IL-6、TXB2蛋白表達（P＜0.05、0.01）；顯著升高p-PLC β3蛋白表達（P＜0.05、0.01）；NETs產(chǎn)生明顯減少，顯著降低P-selectin和CD11b蛋白表達（P＜0.05、0.01）。結(jié)論 Sul-F通過促進p-PLC β3蛋白表達，減少血小板活化，抑制血小板介導(dǎo)的NETs形成，減輕炎癥反應(yīng)，進而改善腦I/R損傷。

Objective To study the mechanism of sodium formononetin-3'-sulphonate (Sul-F) in regulating neutrophil extracellular traps (NETs) to alleviate cerebral ischemia/reperfusion (I/R) injury.Methods SD rats were randomly divided into five groups: control group, model group, arginine aspirin group (AAFI, 200 mg·kg^-1), Sul-F low and high dose groups (20, 40 mg·kg^-1). Except for the control group, the other groups were modeled by middle cerebral artery occlusion, and sacrificed after ischemia for 2 h and reperfusion for 24 h. The drugs were given by ip injection at 0 h and 12 h after reperfusion, and the control group and the model group were given the same volume of 0.9% sodium chloride solution. Neurological function was assessed by the Bederson score. TTC staining was used to determine the cerebral infarction volume. Western blotting was used to detect the expression of phospholipase C-β3(PLC β3) and phosphorylated PLC β3(p-PLC β3) in brain homogenate, and to verify its signaling pathway. The contents of thromboxane B2 (TXB2), interleukin-1β (IL-1β) and interleukin-6 (IL-6) in brain homogenate were detected by enzyme-linked immunosorbent assay. The expression of P-selectin and neutrophil surface adhesion molecule (CD11b) on platelet activation and neutrophil activation was detected by immunofluorescence. H3BB (H3B), myeloperoxidase (MPO) and deoxyribonucleic acid (DNA) were detected by three-color immunofluorescence.Results Compared with the control group, the neurological function score, brain index, cerebral infarction rate, and the protein expressions of IL-1β, IL-6, and TXB2 in brain homogenate of rats in the model group were significantly increased (P<0.01), while the protein expression of p-PLC β3 was significantly decreased (P<0.01), and a large number of NETs were formed. The expressions of P-selectin and CD11b were both increased (P<0.01). Compared with the model group, Sul-F significantly reduced the neurological function score, brain index, cerebral infarction rate, and the protein expressions of IL-1β, IL-6, and TXB2 in brain homogenate (P<0.05, 0.01); Significantly increased the protein expression of p-PLC β3 (P<0.05, 0.01); The production of NETs was significantly reduced, and the protein expressions of P-selectin and CD11b were significantly decreased (P<0.05, 0.01).Conclusion Sul-F improves cerebral I/R injury by promoting the expression of p-PLC β3 protein, inhibiting PLC β3 expression, reducing platelet activation, inhibiting NETs formation, and reducing inflammatory response.

R285.5

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